Biologic Evaluation Of 64Cu & 67Cu As Potential Theranostic Agents For Cancer Radiotherapy

Authors

• Lakshmi Madhusoodanan, University of Alabama at Birmingham

Advisor(s)

Benjamin Larimer

Committee Member(s)

Jonathan McConathy
Natalie Gassman

School

Joint Health Sciences (Interdisciplinary)

Document Type

Thesis

Department (new version)

Joint Health Sciences

Date of Award

9-9-2024

Abstract

Immunotherapy has become a mainstay in the treatment of several types of cancer, with the potential to induce durable remission, albeit in a limited percentage of patients. More recently, radiopharmaceuticals have gained recognition in cancer treatment due to their ability to localize within cancer cells and deliver radiation, reducing adverse effects precisely. Cancer radiotherapy has the potential to synergize with immunotherapy due to its induction of immunogenic cell death (ICD), which results in distinct cellular injuries such as the formation of single-strand breaks (SSBs) and double-stranded breaks (DSBs), disruption of mitochondrial integrity, and the excessive generation of reactive oxygen species (ROS). Currently, there is limited information on the optimal dose, timing, and subcellular localization of radiotherapy to maximize ICD. Copper is an essential micronutrient involved in physiological processes in health and disease conditions. The accumulation of copper in numerous cancers has been observed, and combining radiocopper with immunotherapy has shown improved responses compared to immunotherapy alone. However, the specific mechanism responsible for this enhanced response is still unknown. This study focuses on understanding how beta and Auger radiation delivered by target radiation therapy with 64/67Cu induces cancer ICD. ICD results in distinct cellular injuries such as the formation of SSBs and DSBs, disruption of mitochondrial integrity, and the excessive generation of reactive oxygen species (ROS). Currently there is limited information on the optimal dose, timing, and subcellular localization of radiotherapy to maximize ICD. Therefore, we first examined copper uptake and subcellular localization in MC38 and B16F10 cell lines to understand the role these factors play in the induction of ICD. 64CuCl2, and copper chelated with NOTA (1,4,7-triazacyclononane-1,4,7-triacetic acid) a bifunctional chelator, incubated with MC38 and B16F10 cells allows measurement of cell uptake of free and bound Cu at 1 h, 4 h, and 24 h after dosing. We then evaluated the dose-dependent cell viability and proliferation responses to the radiocopper therapy in these cell lines with clonogenic and cell survival assays. Finally, using the MC38 cell line, we examined ROS levels, mitochondrial damage, and DNA strand breaks post-radiocopper treatment to dissect its ability to induce ICD. ROS and 53BP1 expression, indicating DNA DSB repair, increased after radiocopper exposure. The results indicated a higher intake of radiocopper within the cells, resulting in increased production of ROS, DNA damage at moderate radiation dosage, reduced clonogenic potential, and increased cytotoxic potential as the dosage increased. The differences in response to radiocopper treatment between the two cell lines may be attributed to their respective antioxidant systems. These differences suggest that the effectiveness of radiocopper in inducing ROS and DNA damage may be influenced by the distinct antioxidant capacities of MC38 and B16F10 cells. This study offers valuable insights into the potential biological responses in tumor progression and the underlying cytotoxicity when combining radiocopper treatment with immunotherapy.

ProQuest ID

https://proquest.com/docview/3101886830